Methods and compositions for producing vitamin K dependent proteins
AssigneeThe University of North Carolina at Chapel Hill
InventorsDarrel W. Stafford, Tao Li
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Abstract
The present invention relates to methods and compositions for improving the productivity of recombinant vitamin K dependent protein expression in host cells.
Claims24
An isolated host cell comprising a recombinant nucleic acid coding for a vitamin K epoxide reductase (VKOR), wherein the recombinant nucleic acid coding for the VKOR comprises a nucleotide sequence selected from the group consisting of:
- 1) the nucleotide sequence of SEQ ID NO:9;
- 2) a nucleotide sequence that encodes the amino acid sequence of SEQ ID NO:
- 10;
- 3) a nucleotide sequence having at least 95% identity with the nucleotide sequence of SEQ ID NO:9 and that encodes a VKOR;
- 4) a nucleotide sequence that hybridizes with the complement of the nucleotide sequence of (1) or (2) above under stringent conditions as represented by hybridization conditions of 0.5M NaHPO4, 7% sodium dodecyl sulfate (SDS), 1mM EDTA at 65° C. and wash conditions of 0.1×SSC/0.1% SDS at 68° C. and that encodes a VKOR;
- 5) a nucleotide sequence that encodes an amino acid sequence having at least 95% identity with the amino acid sequence of SEQ ID NO:
- 10 and that encodes a VKOR;
- and
- 6) a nucleotide sequence that differs from the nucleotide sequence of(1), (2), (3), (4) or (5) above due to the degeneracy of the genetic code and that encodes a VKOR,
- and a recombinant nucleic acid coding for a vitamin K dependent protein, wherein both the recombinant VKOR and the recombinant Vitamin K dependent protein are expressed in said host cell.
A cell culture system comprising isolated cells that contain a recombinant nucleic acid coding for a vitamin K epoxide reductase (VKOR), wherein the recombinant nucleic acid coding for the VKOR comprises a nucleotide sequence selected from the group consisting of:
- 1) the nucleotide sequence of SEQ ID NO:9;
- 2) a nucleotide sequence that encodes the amino acid sequence of SEQ ID NO:10;
- 3) a nucleotide sequence having at least 95% identity with the nucleotide sequence of SEQ ID NO:9 and that encodes a VKOR;
- 4) a nucleotide sequence that hybridizes with the complement of the nucleotide sequence of (1) or (2) above under stringent conditions as represented by hybridization conditions of 0.5M NaHPO4, 7% sodium dodecyl sulfate (SDS), 1mM EDTA at 65° C. and wash conditions of 0.1×SSC/0.1% SDS at 68° C. and that encodes a VKOR;
- 5) a nucleotide sequence that encodes an amino acid sequence having at least 95% identity with the amino acid sequence of SEQ ID NO:
- 10 and that encodes a VKOR;
- and
- 6) a nucleotide sequence that differs from the nucleotide sequence of(1), (2), (3), (4) or (5) above due to the degeneracy of the genetic code and that encodes a VKOR,
- and a recombinant nucleic acid coding for a vitamin K dependent protein, wherein both the recombinant VKOR and the recombinant Vitamin K dependent protein are expressed in said cells.
Dependencies are inferred from the claim text.